Immune regulatory oligonucleotide (IRO) compounds to modulate toll-like receptor based immune response

Abstract

The invention provides novel immune regulatory oligonucleotides (IRO) as antagonist of TLRs and methods of use thereof. These IROs have unique sequences that inhibit or suppress TLR-mediated signaling in response to a TLR ligand or TLR agonist. The methods may have use in the prevention and treatment of cancer, an autoimmune disorder, airway inflammation, inflammatory disorders, infectious disease, skin disorders, allergy, asthma or a disease caused by a pathogen.

1. An immune regulatory oligonucleotide (IRO) compound comprising at least two oligonucleotides linked by a non-nucleotide linker at their 3′ ends or by a functionalized sugar or by a functionalized base via a non-nucleotide linker, wherein at least one oligonucleotide has the structure
   5′-N m —N 3 N 2 N 1 CGN 1 N 2 N 3 —N m -3′:
 
 wherein: 
 CG is an oligonucleotide motif that is CpG, C*pG, C*pG* or CpG*, wherein C is cytosine, C* is a pyrimidine nucleotide derivative, G is guanosine, and G* is a purine nucleotide derivative; N 1  is a modified nucleoside that suppresses the activity of the oligonucleotide motif selected from the group consisting of 2′-substituted ribonucleoside, 2′-O-substituted ribonucleoside, 2′-substituted arabinoside, and 2′-O-substituted arabinoside; 
 N 2 -N 3 , at each occurrence, is independently i) a nucleotide, ii) a nucleotide derivative, or iii) a modified nucleoside that suppresses the activity of the oligonucleotide motif selected from the group consisting of 2′-substituted ribonucleoside, 2′-O-substituted ribonucleoside, 2′-substituted arabinoside, and 2′-O-substituted arabinoside; 
 N 1 -N 3 , at each occurrence, is independently i) a nucleotide, or ii) a nucleotide derivative; 
 N n , and N m , at each occurrence, is independently a nucleotide, nucleotide derivative or non-nucleotide linkage; 
 and further provided that the compound contains less than 3 consecutive guanosine nucleotides; 
 wherein the oligonucleotide motif would be immune stimulatory but for the one or more modified nucleosides that suppress the activity of the oligonucleotide motif; 
 wherein m is a number from 0 to about 30; 
 wherein the IRO is an antagonist of an agonist of TLR7, TLR8 and/or TLR9; and 
 provided that the at least two oligonucleotides are not antisense oligonucleotides. 
 
     
     
       2. A pharmaceutical composition comprising an IRO according to  claim 1  and a pharmaceutically acceptable carrier. 
     
     
       3. The IRO according to  claim 1 , wherein the non-nucleotide linker linking the at least two oligonucleotides at their 3′ ends or functionalized base or sugar is Glycerol (1,2,3-Propanetriol), 1,2,4, Butanetriol, 2-(hydroxymethyl)-1,3-propanediol, 2-(hydroxymethyl)1,4-butanediol, 1,3,5-Pentanetriol, 1,1,1-Tris(hydroxymethyl)ethane, 1,1,1-Tris(hydroxymethyl)-nitromethane, 1,1,1-Tris(hydroxymethyl)propane, 1,2,6-Hexanetriol, 3-Methyl-1,3,5-pentanetriol, 1,2,3-Heptanetriol, 2-Amino-2-(hydroxymethyl)-1,3-propanediol, N-[Tris(hydroxymethyl)methyl]acrylamide, cis-1,3,5-Cyclohexanetriol, Cis-1,3,5-Tri(hydroxymethyl)cyclohexane, 3,5-Di(hydroxymethyl)phenol, 1,3,5-Trihydroxyl-benzene, 3,5-Di(hydroxymethyl)benzene, 1,3-Di(hydroxyethoxy)-2-hydroxyl-propane, 1,3-Di(hydroxypropoxy)-2-hydroxyl-propane, 2-Deoxy-D-ribose, 1,2,4-Trihydroxyl-benzene, D-Galactoal, 1,6-anhydro-β-D-Glucose, 1,3,5-Tris(2-hydroxyethyl)-Cyanuric acid, Gallic acid, 3,5,7-Trihydroxyflavone, 4,6-Nitropyrogallol, Ethylene glycol, 1,3-Propanediol, 1,2-Propanediol, 1,4-Butanediol, 1,3-Butanediol, 2,3-Butanediol, 1,4-Butanediol, 1,5-Pentanediol, 2,4-Pentanediol, 1,6-Hexanediol, 1,2-Hexanediol, 1,5-Hexanediol, 2,5-Hexanediol, 1,7-Heptanediol, 1,8-Octanediol, 1,2-Octanediol, 1,9-Nonanediol, 1,12-Dodecanediol, Triethylene glycol, Tetraethylene glycol, 2-(1-Aminopropyl)-1,3-propanediol, or 1,2-Dideoxyribose. 
     
     
       4. The IRO according to  claim 1 , wherein the non-nucleotide linker linking the at least two oligonucleotides at their 3′ ends or functionalized base or sugar is Glycerol (1,2,3-Propanetriol). 
     
     
       5. The IRO according to  claim 1 , wherein the pyrimidine nucleotide derivative is 2-deoxythymidine, 1-(2′-deoxy-α-D-ribofuranosyl)-2-oxo-7-deaza-8-methyl-purine, 2′-dideoxy-5-halocytosine, 2′-dideoxy-5-nitrocytosine, arabinocytidine, T-deoxy-5-hydroxycytidine, 2′-deoxy-N4-alkyl-cytidine, 2′-deoxy-4-thiouridine, or other pyrimidine nucleoside analogs. 
     
     
       6. The IRO according to  claim 1 , wherein the purine nucleotide derivative is 2′-deoxy-7-deazaguanosine, 2′-deoxy-6-thioguanosine, arabinoguanosine, 2′-deoxyinosine, or other purine nucleoside analogs. 
     
     
       7. The IRO according to  claim 1 , wherein at least one of the oligonucleotides has the sequence selected from TCT GA CGTTCT (SEQ ID NO: 86), TCT GA CG 1 TTCT (SEQ ID NC): 87), TCT GA CG 4 TTCT (SEQ ID NO: 88), TCTCT GA CGTT (SEQ ID NO: 89), TCT GU CGTTCT (SEQ ID NO: 93), TCT GU CG 1 TTCT (SEQ ID NO: 94), TCT GA CG 4 TTCT (SEQ ID NO: 95), TCT GA CG 1 TT (SEQ ID NO: 96),  UGU CG 1 TTCT (SEQ ID NO: 100) and  UGA CG 1 TTCT (SEQ ID NO: 101), wherein G 1 =7-deaza, G 4 =araG, and  G ,  A  or  U =2′-OMe. 
     
     
       8. The IRO according to  claim 1 , wherein the IRO is selected from 5′-(TCT GA CGTTCT) 2 X 2 (5′-SEQ ID NO: 86-3′-X 2 -3′-SEQ ID NO: 86-5′), 5′-(TCT GA CG 1 TTCT) 2 X 2 (5′-SEQ ID NO: 87-3′-X 2 -3′-SEQ ID NO: 87-5′), 5′-(TCT GA CG 4 TTCT) 2 X 2 (5′-SEQ ID NO: 88-3′-X 2 -3′-SEQ ID NO: 88-5′), 5′-(TCTCT GA CGTT) 2 X 2 (5′-SEQ ID NO: 89-3′-X 2 -3′-SEQ ID NO: 89-5′), 5′-(TCT GU CGTTCT) 2 X 2 (5′-SEQ ID NO: 93-3′-X 2 -3′-SEQ ID NO: 93-5′), 5′-(TCT GU CG 1 TTCT) 2 X 2 (5″-SEQ ID NO: 94-3′-X 2 -3′-SEQ ID NO: 94-5′), 5′-(TCT GA CG 4 TTCT) 2 ×2(5′-SEQ ID NO: 95-3′-X 2 -3′-SEQ ID NO: 95-5′), 5′-(TCT GA CG 1 TT) 2 X 2 (5′-SEQ ID NO: 96-3′-X 2 -3′-SEQ ID NO: 96-5′), 5′-( UGU CG 1 TTCT) 2 X 2 (5′-SEQ ID NO: 100-3′-X 2 -3′-SEQ ID NO: 100-5′) and 5′-( UGA CG 1 TTCT) 2 X 2 (5′-SEQ ID NO: 101-3′-X 2 -3′-SEQ ID NO: 101-5′), wherein G 1 =7-deaza, G 4 =araG,  G ,  A  or  U =2′-OMe and X 2 =glycerol linker. 
     
     
       9. A pharmaceutical composition comprising an IRO according to  claim 7  and a pharmaceutically acceptable carrier. 
     
     
       10. A pharmaceutical composition comprising an IRO according to  claim 8  and a pharmaceutically acceptable carrier. 
     
     
       11. The IRO according to  claim 1 , wherein N 2  is a 2′-substituted ribonucleoside, 2′-O-substituted ribonucleoside, 2′-substituted arabinoside, or 2′-O-substituted arabinoside. 
     
     
       12. The IRO according to  claim 1 , wherein the 2′-O-substituted ribonucleoside of N 1  is a 2′-OMe-ribonucleoside. 
     
     
       13. The IRO according to  claim 11 , wherein the 2′-O-substituted ribonucleoside is a 2′-OMe-ribonucleoside. 
     
     
       14. The IRO according to  claim 1 , wherein the non-nucleotide linker may be attached to the 3′ hydroxyl. 
     
     
       15. The IRO according to  claim 1 , wherein CG is an oligonucleotide motif that is C*pG, C*pG* or CpG*.